Question:
What are the cell calling updates in the Cell Ranger v7.1 release? How do the changes affect my results in Gene Expression assays?
Answer:
For every new release, we have a high level overview of changes in our release notes pages. However in most cases, the changes in the analysis results are data dependent. In this article, we use a few public datasets to illustrate the cell calling changes and its impact on Web Summary metrics in Cell Ranger v7.1 compared to Cell Ranger v7.0 for Single Cell Gene Expression products.
Cell Calling Changes:
In Cell Ranger v7.1, we made a minor change in the cell calling algorithm. We updated the range of barcodes to be used for expect-cells auto estimation. In the previous releases, the upper range was ~260k. In Cell Ranger v7.1, we restricted the upper range to:
- For Single Cell Gene Expression, this value is 45,000 for both standard and high-throughput assay.
- For Single Cell Gene Expression Flex (Fixed RNA Profiling), this value is 45,000, or number of probe barcodes × 22,500, whichever is larger.
We made this improvement with the goal of achieving more accurate estimation of expect-cells parameter, and in turn more reliable cell calling results. Through the hundreds of samples tested, the effect of this change on cell number is very minor (a change of ~0.1% when there is a change, usually positive). Please see a few examples below using our public datasets.
Single Cell Gene Expression:
We analyzed our 10k PBMC dataset using Cell Ranger v7.0.1 and v7.1.0, and the table below summarizes the main metrics:
| Web Summary metrics | Cell Ranger v7.0.1 multi |
Cell Ranger v7.1.0 multi |
| estimated number of cells | 11,996 | 11,999 |
| mean reads per cell in library | 41,379 | 41,369 |
| median reads per cell in sample | 33,882 | 33,876 |
| median UMI counts per cell | 10,180 | 10,178 |
| median genes per cell | 3,267 | 3,266 |
We observed a minor increase of cell count (3 cells), which in-turn impacted other metrics. However, the overall changes are very minor and mostly negligible.
Single Cell Gene Expression Flex:
We analyzed the 40k 4-plex tumor cells using Cell Ranger v7.0.1 and Cell Ranger v7.1.0, and table below shows the comparisons of the main metrics:
|
metrics category |
metrics |
Cell Ranger v7.0.1 multi |
Cell Ranger v7.1.0 multi |
| Cell Statistics |
estimated number of cells |
26,870 | 26,922 |
| metrics per probe barcode (cells) | Bladder_BC1 | 8,371 (31.15%) | 8,372 (31.10%) |
| Breast_BC2 | 5,880 (21.88%) | 5,880 (21.84%) | |
| Kidney_BC4 | 6,861 (25.53%) | 6,857 (25.47%) | |
| Ovary_BC3 | 5,758 (21.43%) | 5,813 (21.59%) |
The changes of cell counts are very minor.
In summary, cell calling changes in Cell Ranger v7.1 should have minimal effect on your results on Gene Expression products.
For more guidance on whether to rerun data analysis using the latest version, please see this article: My samples are analyzed with Cell Ranger v7.0. Should I reanalyze them using the latest Cell Ranger 7.1?
Products: Single Cell Gene Expression, Single Cell Gene Expression Flex